The long-term objective of this project is a combined biochemical and genetic analysis of the hormonal regulation of cell membrane characteristics relevant to neoplasia. These studies will focus on the glucocorticoid and cyclic nucleotide regulation of the serine protease, plasminogen activator in HTC rat hepatoma cells in culture. HTC cells are a suitable experimental model for studying the hormonal regulation of membrane properties because of their well-characterized hormonal responsiveness and the availability of specific variant cell lines altered in hormone responsiveness. The glucocorticoid dexamethasone rapidly inhibits plasminogen activator activity, whereas cyclic nucleotides dramatically stimulate activator activity; paradoxically, dexamethasone enhances this stimulation. We have demonstrated that the glucocorticoid inhibition is secondary to the induction of a 50 kilodalton inhibitor specific for plasminogen activation rather than to a decrease in the amount of activator. We have isolated a unique set of variant HTC cell lines selectively resistant to the dexamethasone inhibition in which glucocorticoids fail to induce the inhibitor. Using specific antibodies, we have shown that HTC plasminogen activator is of the tissue-activator type. We have established a radioimmunoassay with which to analyze the regulation of plasminogen activator by glucocorticoids and cyclic nucleotides. We are planning to purify the glucocorticoid-induced inhibitor, prepare antibodies to it, and study its hormonal regulation at the molecular level. Limited proteolysis mediated by the plasminogen activator-plasmin cascade plays a significant role in the behavior of transformed cells as well as in many normal processes. These studies should increase our understanding of the hormonal regulation of this important cell membrane property and of the membrane biology of neoplastic cells. (A)